?d T cells constitute an important component of the immune response against infectious agents and cancerous transformations, yet the biochemical mechanisms by which they detect antigen through their somatically recombined T cell receptor (TCR) remain unclear. Unlike a[unreadable]TCRs, which are restricted to recognizing antigens in the context of Major Histocompatibility Complex (MHC) molecules, ?d TCRs can recognize a diversity of ligands ranging from self MHC to intact, unprocessed, viral glycoproteins. While the ?d TCR is structurally similar to the a[unreadable]TCR, and both use components of the CD3 signaling complex for signal transduction, it is clear ?d TCRs bind antigen in a distinct and divergent way. The lack of concrete biochemical characterization of these receptors and their interactions with ligands has hindered a true understanding of the role of these receptors in ?d T cell activation. Thus, the long-term goal of this proposal is to understand the biochemical and structural mechanisms of ?d TCR engagement and how this binding event discriminates between healthy and unhealthy tissue to initiate T cell activation. We plan to study these questions using two parallel strategies that are directly complementary, yet unique. The first is a structure/function-based approach combining biochemistry, structural biology and cell-based functional assays to determine the specific molecular details that govern ?d TCR recognition of defined ligands. We will focus our biochemical, biophysical and structural studies on two receptor/ligand interactions: ?d TCRs specific for 1) the murine MHC T107T22 molecules and 2) the human MHC CD1c molecules. The second approach involves using a cross-species sequence-based analysis to understand the selective, evolutionary mechanisms that shape the ?d TCR V, D and J gene repertoire. This approach will define what types of selective pressures (diversifying, neutral or purifying) have governed the evolution of these genes in humans and non-human primates, indicating the nature of the ligands to which they bind. These approaches, together, will help to elucidate the molecular recognition principles of ?d TCR interactions, and determine if ?d TCRs recognize ligands in a convergent manner, such as seen in a[unreadable] TCR/MHCp interactions, or whether there are a diversity of recognition solutions, unique to each ?d T cell population and particular ligand.